单细胞 RNA 测序分析鉴定出铁死亡软骨细胞簇，并揭示 TRPV1 是骨关节炎中的一种抗铁死亡靶点。

BACKGROUND: Osteoarthritis (OA) is the most common degenerative joint disease primarily characterized by cartilage destruction. The aim of this study was to investigate the role, molecular characteristics and potential therapeutic target of chondrocyte ferroptosis in the pathogenesis of OA. METHODS: The expression of ferroptotic hallmarks (iron and lipid peroxidation accumulation, glutathione deletion) were analyzed in paired intact and damaged cartilages from OA patients. Single cell RNA sequencing (scRNA-seq) analysis was performed on 17,638 chondrocytes to verify the presence, investigate the molecular signatures and unveil the potential therapeutic target of ferroptotic chondrocyte cluster in human OA cartilages. Destabilization of medial meniscus (DMM)-induced OA model and tert-butyl hydroperoxide (TBHP)-treated primary mouse chondrocytes and human cartilage explants were used to evaluate the protective effect of pharmacologically activated transient receptor potential vanilloid 1 (TRPV1). The downstream molecular mechanisms of TRPV1 was further investigated in glutathione peroxidase 4 (Gpx4) heterozygous genetic deletion mice (Gpx4). FINDINGS: The concentrations of iron and lipid peroxidation and the expression of ferroptotic drivers in the damaged areas of human OA cartilages were significantly higher than those in the intact cartilage. scRNA-seq analysis revealed a chondrocyte cluster characterized by preferentially expressed ferroptotic hallmarks and genes, namely ferroptotic chondrocyte cluster. Comprehensive gene set variation analysis revealed TRPV1 as an anti-ferroptotic target in human OA cartilage. Pharmacological activation of TRPV1 significantly abrogated cartilage degeneration by protecting chondrocytes from ferroptosis. Mechanistically, TRPV1 promoted the expression of GPX4, and its anti-ferroptotic role was largely mitigated in the OA model of Gpx4 mice. INTERPRETATION: TRPV1 activation protects chondrocytes from ferroptosis and ameliorates OA progression by upregulating GPX4. FUNDING: National Key R&D Program of China (2018YFC1105904), Key Program of NSFC (81730067), National Science Foundation of China (81772335, 81941009, 81802196), Natural Science Foundation of Jiangsu Province, China (BK20180127), Jiangsu Provincial Key Medical Talent Foundation, Six Talent Peaks Project of Jiangsu Province (WSW-079).

背景：骨关节炎（OA）是最常见的退行性关节疾病，主要特征为软骨破坏。本研究旨在探讨软骨细胞铁死亡在 OA 发病机制中的作用、分子特征和潜在治疗靶点。

方法：分析 OA 患者配对完整和受损软骨中的铁死亡特征标志物（铁和脂质过氧化积累、谷胱甘肽缺失）。对 17638 个软骨细胞进行单细胞 RNA 测序（scRNA-seq）分析，以验证铁死亡软骨细胞簇的存在，研究其分子特征，并揭示人 OA 软骨中铁死亡软骨细胞簇的潜在治疗靶点。采用内侧半月板不稳定（DMM）诱导的 OA 模型和叔丁基过氧化氢（TBHP）处理的原代小鼠软骨细胞和人软骨外植体来评估药理学激活瞬时受体电位香草酸 1（TRPV1）的保护作用。进一步在谷胱甘肽过氧化物酶 4（Gpx4）杂合基因缺失小鼠（Gpx4）中研究 TRPV1 的下游分子机制。

结果：人 OA 软骨损伤区的铁和脂质过氧化浓度以及铁死亡驱动因子的表达明显高于完整软骨。scRNA-seq 分析显示，一个软骨细胞簇的特征是优先表达铁死亡特征标志物和基因，即铁死亡软骨细胞簇。综合基因集变异分析显示 TRPV1 是人类 OA 软骨中的一种抗铁死亡靶点。TRPV1 的药理学激活通过保护软骨细胞免受铁死亡显著减轻了软骨退变。在 Gpx4 小鼠的 OA 模型中，TRPV1 通过促进 GPX4 的表达，其抗铁死亡作用在很大程度上得到缓解。

结论：TRPV1 的激活通过上调 GPX4 来保护软骨细胞免受铁死亡，从而改善 OA 的进展。

资助：国家重点研发计划（2018YFC1105904）、国家自然科学基金重点项目（81730067）、国家自然科学基金（81772335、81941009、81802196）、江苏省自然科学基金、江苏省医学重点人才项目、江苏省六大人才高峰项目（WSW-079）。